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Yields of tyrothricin in excess of 2 grams per liter of medium were obtained through the growth of Bacillus brevis (B.G.) in shallow layers of medium. Maximum yields were found after 10 or 16 days of incubation at about 35°C. with the medium disposed in 11-mm. layers. Complex sources of nitrogen, such as Bacto tryptone, acid hydrolyzate of casein, corn-steep liquor, tryptic digest of soybean meal, and press juice concentrates from waste asparagus butts, proved most suitable; relatively simple substances, such as glutamic acid, asparagine, or ammonium sulfate plus citric or malic acid, proved moderately effective in the presence of 0.2% Bacto tryptone.About 3 to 5% of a fermentable carbon compound, such as glucose, mannitol, or glycerol, was necessary for best yields; fructose, sucrose, lactose, and maltose proved much less effective. Requirements for calcium, magnesium, and manganese were demonstrated.

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